Question

What are the components of agarose gel electrophoresis set up?

Answer 1

Agarose gel electrophoresis set up:

Gels are described in terms of percents: 0.7%, 0.8%, 1%, and 1.2% are pretty common gel percentages. The gel percentage depends on a few things: such as size fragment

The percentage measurement is a weight/volume thing. For example, 1% gel would be 1g agarose in 100mL TAE. TAE is Tris-Acetate-EDTA; it’s a buffer and we make gels with TAE and run them in TAE buffer. .

We have gel boxes and casting trays that vary in size. The volume of gel that are needed to make will depend on the size of the casting tray. For the smallest gel trays, 30-40mL is a convenient volume. The wells of the gel are made by inserting a comb into the slots in the tray, and as the agarose hardens around the comb, wells are formed. The thicker the gel, the deeper the wells will be.

To make a gel, first we have to figure out what volume we want. Then pour water into the tray and when the wells look deep enough, record the volume and make gel using that volume.

Then figure out what mass of agarose to use for the percentage gel we want. For example, if we are making a 30mL gel that you want to be 0.8%, the amount of agarose to use is 0.24g. Measure out the agarose using wax weighing paper. Put the agarose in an Erlenmeyer flask. Measure out the correct volume of TAE using a graduated cylinder. Swirl the contents of the flask and cover the top with a paper towel.

Microwave the gel until it melt completely. Let it cool on the benchtop for five or so minutes.

Next add ethidium bromide - chemical that intercalates with DNA and makes it visible under UV light. The amount of EtBr to add is as follows: of a 0.5mg/mL stock solution, add 1/1000 to the gel. For example,In 30mL gel, add 30°L of EtBr.

The casting trays have rubber stuff around the edges that makes a tight seal when you put them in the gel boxes sideways. They also have notches where we can insert a comb. The size comb to use depends on the width we want our wells to be. Swirl the flask immediately before pouring the gel into the tray to make sure it’s mostly all at the same temperature; otherwise the gel will harden. It will take about 20 minutes for a small gel to harden enough to be used, longer for bigger gels.

Hence, components of agarose gel electrophoresis are: Agarose gel,TAE buffer, Etbr-ethidium bromide,Erlenmeyer flak,weight machine,gel boxes, casting trays, wax weighing papers.